Victoria University, Melbourne, VIC, Australia.
J Ind Microbiol Biotechnol. 2010 Feb;37(2):139-49. doi: 10.1007/s10295-009-0655-3. Epub 2009 Nov 10.
Saccharomyces spp. are widely used for ethanologenic fermentations, however yeast metabolic rate and viability decrease as ethanol accumulates during fermentation, compromising ethanol yield. Improving ethanol tolerance in yeast should, therefore, reduce the impact of ethanol toxicity on fermentation performance. The purpose of the current work was to generate and characterise ethanol-tolerant yeast mutants by subjecting mutagenised and non-mutagenised populations of Saccharomyces cerevisiae W303-1A to adaptive evolution using ethanol stress as a selection pressure. Mutants CM1 (chemically mutagenised) and SM1 (spontaneous) had increased acclimation and growth rates when cultivated in sub-lethal ethanol concentrations, and their survivability in lethal ethanol concentrations was considerably improved compared with the parent strain. The mutants utilised glucose at a higher rate than the parent in the presence of ethanol and an initial glucose concentration of 20 g l(-1). At a glucose concentration of 100 g l(-1), SM1 had the highest glucose utilisation rate in the presence or absence of ethanol. The mutants produced substantially more glycerol than the parent and, although acetate was only detectable in ethanol-stressed cultures, both mutants produced more acetate than the parent. It is suggested that the increased ethanol tolerance of the mutants is due to their elevated glycerol production rates and the potential of this to increase the ratio of oxidised and reduced forms of nicotinamide adenine dinucleotide (NAD(+)/NADH) in an ethanol-compromised cell, stimulating glycolytic activity.
酿酒酵母被广泛用于乙醇发酵，但随着发酵过程中乙醇的积累，酵母的代谢率和活力会下降，从而降低乙醇的产量。因此，提高酵母的乙醇耐受性应该可以降低乙醇毒性对发酵性能的影响。本研究的目的是通过以乙醇胁迫作为选择压力，对酿酒酵母 W303-1A 的诱变和非诱变群体进行适应性进化，从而产生并鉴定具有乙醇耐受性的酵母突变体。突变体 CM1（化学诱变）和 SM1（自发突变）在亚致死乙醇浓度下培养时，具有更高的适应和生长速率，与亲本菌株相比，其在致死乙醇浓度下的存活率有了显著提高。与亲本菌株相比，在存在乙醇和初始葡萄糖浓度为 20 g l(-1)时，突变体在乙醇存在的情况下能够以更高的速率利用葡萄糖。在 100 g l(-1)葡萄糖浓度下，SM1 在存在或不存在乙醇的情况下具有最高的葡萄糖利用率。突变体产生的甘油量明显多于亲本，尽管只有在乙醇胁迫培养物中才能检测到乙酸，但与亲本相比，两个突变体都产生了更多的乙酸。推测突变体的乙醇耐受性增加是由于其甘油生成率的提高，以及这可能增加了在乙醇胁迫下细胞中氧化型和还原型烟酰胺腺嘌呤二核苷酸（NAD(+)/NADH）的比例，从而刺激糖酵解活性。
